We plan to isolate lysosomes from cultured normal and cystinotic fibroblasts to allow us to study their mediated uptake and release of amino acids. We hope thereby to identify, characterize, and relate transport systems mediating the movement of cystine, glutamate, asparate, cysteine and other amino acids, both inwardly and outwardly, also using metabolism-resistant analogs in strategies of inhibition analysis and trans stimulation already developed for intact fibroblasts and other cells. We will vary the pH at each membrane surface to modify the form in which an amino acid is presented; we will modify the amino acid structure itself, also the ionic composition of the two aqueous phases, in our effort to attain these descriptions. Our goal is not merely to help complete and confirm the transport defect of cystinosis, but to explore more widely the role of amino acid transport in the regulation of lysosomal function. Other health consequences of defective lysosomal transport besides nephropathic cystinosis may be uncovered. In this collaboration a biochemical laboratory with a substantial motivation and experience in studying the physiological (biophysical, cell biological, nutritional) problems of membrane transport will be joining hands with a pediatric genetics research laboratory representing a parallel motivation and experience in cystinosis and other inborn diseases of amino acid metabolism.